Human osteoarthritis (OA) is a progressive joint disease characterized by degradation of articular cartilage. Other key features normally occurring during OA development include thickening of the subchondral bone, formation of osteophyte, variable degrees of inflammation of the synovium, degeneration of ligaments and the menisci, and hypertrophy of the joint capsule. However, the consequence and relationship among these events are currently unknown. TGF? signaling plays a critical role in controlling chondrocyte differentiation. It has been reported that inhibition of TGF? signaling causes severe OA-like phenotype in mice. Recent genetic studies also demonstrated that mutations of the Smad3 gene are associated with high incidence of hip and knee OA in patients. However, the key downstream target genes of TGF? signaling in articular chondrocytes during OA development remains to be determined. Our preliminary studies demonstrated that chondrocyte-specific deletion of the type II TGF? receptor gene (Tgfbr2) at postnatal/adult stages lead to development of a severe OA-like phenotype. Tgfbr2 conditional knockout (Tgfbr2Col2ER) mice display the most important features of OA. In these mice Runx2 and Atf4 expression was significantly increased. We further demonstrated that inhibition of TGF? signaling up regulates Mmp13 and Adamts5 expression. We generated Tgfbr2/Mmp13 and Tgfbr2/Adamts5 double knockout mice and demonstrated that deletion of either the Mmp13 or Adamts5 gene in the Tgfbr2Col2ER background significantly reversed the majority of the OA features observed in Tgfbr2Col2ER mice. Based on these observations, we hypothesize that Runx2, with ATF4, plays a central role in activation of Mmp13 and Adamts5 expression (MMP13 and ADAMTS5 are two key enzymes in cartilage matrix degradation) and consequently leading to OA development in Tgfbr2Col2ER mice. In the proposed studies we will determine the signaling mechanism of the TGF? inhibition->Runx2/ATF4->Mmp13/Adamts5 pathway in articular chondrocytes using comprehensive molecular and genetic approaches. We have proposed three specific aims to test our hypothesis. In Specific Aim 1: we will determine the role of TGF? signaling in OA development in adult mice. Our hypothesis is that deletion of the Tgfbr2 gene in articular chondrocytes of adult mice leads to OA-like morphological changes and pain-related behavioral changes in adult Tgfbr2Col2ER mice. In Specific Aim 2: we will determine the role of Mmp13 in Tgfbr2Col2ER-induced OA phenotype. Our hypothesis is that TGF? signaling inhibition-induced Mmp13 expression is mediated by transcription factors Runx2 and ATF4. In Specific Aim 3: we will determine the role of Adamts5 in Tgfbr2Col2ER-induced OA phenotype. Our hypothesis is that deletion of Adamts5/Mmp13 genes in Tgfbr2Col2ER mice will have better protective effect than deletion of Mmp13 and Adamts5 alone on Tgfbr2Col2ER-induced articular cartilage degradation. Our proposed studies will provide novel insights into the molecular mechanisms of OA development.